Gibbon ape leukemia virus receptor functions of type III phosphate transporters from CHOK1 cells are disrupted by two distinct mechanisms

The Chinese hamster cell lines E36 and CHOK1 dramatically differ in suscept ibility to amphotropic murine leukemia virus (A-MuLV) and gibbon ape leukem ia virus (GALV); E36 cells are highly susceptible to both viruses, CHOK1 ce lls are not. We have previously shown that GALV can infect E36 cells by usi ng both its own receptor, HaPit1, and the A-MuLV receptor, HaPit2. Given th at the two cell lines are from the same species, the loss of function of bo th of these receptors in CHOK1 cells is surprising. Other studies have show n that CHOK1 cells secrete proteins that block A-MuLV entry into CHOK1 as w ell as E36, suggesting the two A-MuLV receptors are functionally identical. However, CHOK1 conditioned medium does not block GALV entry into E36, indi cating the secreted inhibitors do not block HaPit1. HaPit1 and ChoPit1 ther efore differ as receptors for GALV; ChoPit1 is either inactivated by secret ed factors or intrinsically nonfunctional. To determine why GALV cannot inf ect CHOK1, we cloned and sequenced ChoPit1 and ChoPit2, ChoPit2 is almost i dentical to HaPit2, which explains why CHOK1 conditioned medium blocks A-Mu LV entry via both receptors, Although ChoPit1 and HaPit1 are 91% identical, a notable difference is at position 550 in the fourth extracellular region , shown by several studies to be crucial for GALV infection. Pit1 and HaPit 1 have aspartate at 550, whereas ChoPit1 has threonine at this position. We assessed the significance of this difference fur GALV infection by replaci ng the aspartate 550 in Pit1 with threonine, This single substitution rende red Pit1 nonfunctional fur GALV and suggests that threonine at 550 inactiva tes ChoPit1 as a GALV receptor. Whether native ChoPit1 functions for GALV w as determined by interference assays using Lec8, a glycosylation-deficient derivative of CHOK1 that is susceptible to both viruses and that has the sa me receptors as CHOK1. Unlike with E36, GALV and A-MuLV exhibited reciproca l interference when infecting Lec8, suggesting that they use the same recep tor. We conclude both viruses can use ChoPit2 in the absence of the inhibit ors secreted by CHOK1 and ChoPit1 is nonfunctional.